This research program is designed to isolate and purify biologically active Proinsulin mRNA. This study will be carried out in the following manner: (a) Isolation of crude polysomal RNA from a suitable tissue synthesizing insulin; (b) Initial purification of this RNA by oligo (dt) cellulose chromatography followed by subsequent purification steps using sucrose gradient fractionation; (c) Translation of the purified mRNA in a cell-free system derived from ascites tumor cell and wheat germ; (d) Attempts at identifying the published methods for protein and peptide analyses. Another aim is to quantitate the levels of proinsulin mRNA in isolated islets using the radioactive DNA transcript of proinsulin mRNA and sensitive hybridization techniques. The radioactive DNA will be prepared from purified mRNA with the reverse transcriptase purified from avian myeloblastosis virus. The question of how various physiological effectors stimulate proinsulin synthesis, for example, can then be examined. It is believed that having in hand purified proinsulin mRNA will provide insight as to (a) the molecular mechanisms by which a select class of effector molecules stimulate insulin synthesis and (b) the possible involvement of such effects in disease. BIBLIOGRAPHIC REFERENCES: Permutt, M. A., Biesbroeck, J., Chyn, R., Biome, I., Szczesna, E. and McWilliams, D. Proceedings of the Ciba Symposium, 41, 97-116 (1976). Szczesna, E. and Boime, I., Proceedings National Academy of Science, 73, 1179 (1976).